Effect of new strains of rotaviruses on PD1 and PD-L1 expression on peripheral blood T cells.
Authors: Anastasia O. Sitkovskaya, Oleg I. Kit, Svetlana Yu Filippova, Irina V. Mezhevova, Nadezhda V. Gnennaya, Tatiana V. Shamova, Sofia V. Timofeeva, Inna A. Novikova, Elena Yu Zlatnik, Elena S. Bondarenko, Elena P. Kolpakova, Larisa N. Vashchenko, Emma E. Kechedzhieva, Irina R. Dashkova, Tatiana V. Ausheva, Elena А. Andreiko
Journal: Journal of Clinical Oncology (2022)

e14539 Background: Promising oncolytic viruses are evaluated, as a rule, by their direct cytotoxic effect on cancer cells. However, the positive effect of cancer virotherapy may be associated with an effect on the immune system, in particular, on the expression of PD-1 and PD-L1 on lymphocytes. The aim of the study was to evaluate the effect of unclassified non-pathogenic strains of rotaviruses RVK100 and RVK228 on the expression of PD-1 and PD-L1 on peripheral blood T-cells of breast cancer patients. Methods: PBMC were cultured at 10 6 cells/ml cell density in RPMI 1640 medium (Gibco, USA) without the addition of serum at 37°C in an atmosphere containing 5.0% CO 2 . Four experimental variants were established - 1) negative control without the addition of viruses, 2) positive control of activation with phytohemagglutinin (PHA) addition, 3) addition of RVK100 strain in concentration 10 7 particles/ml, 4) addition of RVK228 strain in concentration 10 7 particles/ml. After 24 and 72 hours of incubation, the expression of PD-1 (CD279) and PD-L1 (CD274) on T cells was determined by flow cytometry. Results: After 24 hours of cultivation, an increase in PD-1 expression on CD4+ cells was observed in samples with addition of PHA - 40.5%, RVK100 - 42.3% and RVK228 - 37.5% compared with the control (18.1%). A similar, although less pronounced increase in PD-1 expression, was observed on CD8+ cells: PHA - 41.7%, RVK100 - 46.4%, RVK228 - 42.6% compared with 27.7% in the control. Expression of PD-L1 on CD4+ cells increased to 67.0% and 58.6% in samples with addition of RVK100 and RVK228 strains, respectively, while under the influence of PHA it increased to 75.1% compared with the control (44.8%). A similar trend was also found on CD8+ cells (control - 46.2%, RVK100 - 63.4%, RVK228 - 58.4%, PHA - 52.8%). After 72 hours, an increase in PD-1 expression on CD4+ cells was observed only in the control (up to 41.2%), while in the experimental variant with RVK100 addition there was a 2-fold decrease (up to 21.6%) and no significant changes were found in samples with addition of PHA and RVK228 compared to 24 h. At the same time, cultivation with RVK100 caused a decrease in PD-1 expression on CD8+ cells by 2.7 times (up to 17.4%) compared with 24 h, without significant changes in other samples (control - 33.1%, PHA - 48.1%, RVK228 - 38.4%). The expression of PD-L1 on CD4+ cells generally remained unchanged compared to 24 h, while proportion of CD8+CD279+ cells increased in all variants and reached 67-79% the in experimental and control samples. Conclusions: Both strains, like the nonspecific T-mitogen PHA, stimulated the expression of immune checkpoint receptors PD-1 and PD-L1 on T-helpers and CTLs after 24 hours of cultivation. After 72 hours of cultivation, RVK100, in contrast to RVK228, was able to reduce the expression of PD-1 on these cells.